Understanding the structures of the toxic oligomers formed by the ?-amyloid peptide A? is paramount for understanding the molecular basis of Alzheimer's disease, and for developing effective diagnostic tools and therapies. This proposal seeks to generate and study a new class of monoclonal antibodies (mAbs) against conformationally and structurally defined oligomers derived from A?. The Nowick laboratory has pioneered X- ray crystallographic structure determination of oligomers formed by macrocyclic ?-hairpin peptide mimics that contain sequences from central and C-terminal regions of A?. They have discovered that these A? ?-hairpin mimics assemble to form triangular trimers that further assemble to form high-order oligomers such as hexamers and dodecamers. To better understand and study the biological significance of the triangular trimers, they have created covalently stabilized variants of the triangular trimers and have shown that these triangular trimers mimic some of the biological and biophysical properties of oligomers formed by full-length A?. The Nowick laboratory has also generated polyclonal antibodies against three different stabilized triangular trimers, and have discovered that these polyclonal antibodies react with oligomers formed in vitro by recombinantly expressed A?, but do not recognize A? monomers or fibrils. Western blot analysis has revealed that these polyclonal antibodies recognize A? oligomers in protein extracts from Alzheimer's disease brains. Immunohistochemistry studies in human and transgenic mouse brain tissue sections have shown that these polyclonal antibodies stain specific extracellular and intracellular features in the brains, and do not stain extracellular plaques. This proposal aims to build on the discoveries made with the polyclonal antibodies. The broad overarching goals are to generate monoclonal antibodies (mAbs) and additional polyclonal antibodies against the triangular trimers and other oligomers and examine their reactivity with A? oligomers, determine the structural basis of antibody specificity for the triangular trimers, and explore the therapeutic potential of the mAbs. To achieve these goals the research team will collaborate with the Lai laboratory to isolate mAbs from rabbits and mice against the stabilized triangular trimers. The research team will also isolate human mAbs from human peripheral blood mononuclear cells. They will characterize the reactivity of the mAbs in vitro with oligomers, monomers, and fibrils of recombinantly expressed A?, and with A? oligomers in protein extracts and brain tissue slices from human Alzheimer's disease brains and brains from Alzheimer's disease transgenic mice. The research team will elucidate the X-ray crystallographic structures of the antigen binding fragments (Fab) complexed with the triangular trimers and oligomers of full-length A?. They will explore the therapeutic potential of the mAbs by investigating if passive immunization with candidate mAbs is beneficial to Alzheimer's disease transgenic mice.